CN103393628A - Application of muscarinic receptor 3 in treating prostatic cancer - Google Patents
Application of muscarinic receptor 3 in treating prostatic cancer Download PDFInfo
- Publication number
- CN103393628A CN103393628A CN2013103022648A CN201310302264A CN103393628A CN 103393628 A CN103393628 A CN 103393628A CN 2013103022648 A CN2013103022648 A CN 2013103022648A CN 201310302264 A CN201310302264 A CN 201310302264A CN 103393628 A CN103393628 A CN 103393628A
- Authority
- CN
- China
- Prior art keywords
- prostate
- tumor
- cell
- chrm3
- chr
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention provides an application of a muscarinic receptor 3 as a drug target in treating prostatic cancer. The application has the advantages that high expression of CHRM3 (Muscarinic Acetylcholine Receptor 3) in tumor of prostate is discovered for the first time, and CHRM3 is activated to promote proliferation and migration of tumor of prostate and is inhibited as the drug target to inhibit growth and migration of tumor of prostate. CHRM3 is inhibited as an important clinical therapeutic target for treating prostatic cancer, so that huge social and economic benefits are brought.
Description
Technical field
The present invention relates to the clinical practice field, specifically, is the application of M-ChR 3 in prostate cancer therapy.
Background technology
Carcinoma of prostate is the modal malignant tumor of male, and 95% betides old people more than 60 years old.Along with aged tendency of population, China has ten thousand carcinoma of prostate of 7-8 to increase case newly every year, and sickness rate, with the speed increment of annual 25%-30%, becomes the fastest malignant tumor of sickness rate increase.Relevant research prompting, if having no idea to control, after 10 years, patients with prostate cancer likely reaches in 100,000 people 60 to 70.Undoubtedly, carcinoma of prostate becomes the major disease of harm elderly men health day by day.
Three kinds of classic treatment methods that clinically carcinoma of prostate adopted at present (radical prostatectomy, antiandrogen treatment and radiotherapy) although very effective at the disease initial stage, all tumor cells that can not the kill tumor tissue.Patient is after accepting these academic therapies, As time goes on, residual, the insensitive tumor cell of academic therapy is become to tumor, migration, propagation in the future, thereby causes Preventive and the deterioration of carcinoma of prostate, finally causes patient death.The domestic research to carcinoma of prostate at present is less, often is confined to the expression of some specific gene.For many years, but the anti-academic therapy of carcinoma of prostate and the mechanism that worsen to shift it be not immediately clear in external research.Drug therapy clinically, except the antiandrogen therapy, there is no other active drug.Mostly inactivation, Growth factors and receptors, and the research of regulating cell apoptosis of the activation of proto-oncogene and antioncogene is paid attention in basic research.But up to the present, not yet develop the medicine of especially effectively treating carcinoma of prostate.
Summary of the invention
The objective of the invention is, for deficiency of the prior art, provides the application of M-ChR 3.
For achieving the above object, the technical scheme that the present invention takes is: the application of M-ChR 3 in the target agent of preparation treatment carcinoma of prostate.
The target agent of described treatment carcinoma of prostate is usingd M-ChR 3 as drug targets.
Second purpose of the present invention is that M-ChR 3 application in the medicine of screening treatment carcinoma of prostate as target is provided.
The 3rd purpose of the present invention is to provide the application of M-ChR 3, as the drug targets for the treatment of carcinoma of prostate medicine.
The 4th purpose of the present invention is that the application of M-ChR 3 receptor antagonists in the medicine of preparation treatment carcinoma of prostate is provided.
Described M-ChR 3 receptor antagonists suppress propagation and the migration of prostate tumor cells.
Described M-ChR 3 receptor antagonists are Tolterodine, P-F-HHSiD, Oxybutynin, Darifenacin, 4-DAMP.
Described M-ChR 3 receptor antagonists are Darifenacin.
The invention has the advantages that:
The present invention finds CHRM3 high expressed in tumor of prostate first, and the activation of CHRM3 can promote propagation and the migration of tumor of prostate, suppresses CHRM3 and can be used as growth and the migration that drug targets suppresses tumor of prostate.The clinical treatment target spot that the inhibition of CHRM3 is important as prostate treatment, can bring huge social and economical benefits.
The accompanying drawing explanation
Figure 1A detects the expression of CHRM3 in tissue array by SABC, more paired tumor (Cancer) and adjacent hyperplastic tissue (BPH), by the optical density of Image pro plus software analysis CHRM3 dyeing, by t assay p=0.003.
Figure 1B detects the expression of the mRNA of CHRM3 in cell line by the method for RT-PCR, wherein PNT1B and BPH1 are non-tumor cell system, and the cell line that LNCaP and CP3-AR are the androgen sensitivity.DU145, C4-2B, 22Rv1 and PC3 are the non-sensitive cell line of androgen.This result shows that CHRM3 expresses and raises in part prostate tumor cells system
.
Fig. 2 A figure is the MTS(cell proliferation detecting kit) detect the picture of cell proliferation, and the carbachol(carbachol) be M3(M-ChR 3) stable activator, specifically see experimental technique.Wherein * means p<0.05, and * * means p<0.01, and * * * means p<0.001; B figure dyes at DU145 and the low transit cell of expressing of these two CHRM3 of LNCaP the virus that CHRM3 crosses expression, sets up stable cell line, then by the MTS method, detects cell proliferation.
Fig. 3 detects the impact of CHRM3 inactivation for prostate tumor cells propagation by EDU, above two by the selective antagonist darifenacin(darifenacin of M3 receptor, (M3) M-ChR blocade), wherein * means p<0.05, * means p<0.01, and * * * means p<0.001.
Fig. 4 A is the variation of the nude mice by subcutaneous tumor model volume of PC3 cell and 22Rv1 cell, and abscissa is natural law, and vertical coordinate is the volume of tumor.Wherein * means p<0.05, and * * means that p<0.01 * * * means p<0.001.B is the variation of the nude mice by subcutaneous tumor model tumor weight of PC3 cell and 22Rv1 cell, and abscissa is natural law, and vertical coordinate is the volume of tumor.Wherein * means p<0.05, and * * means that p<0.01 * * * means p<0.001.
Fig. 5 A crosses expression CHRM3 in the PNT1B cell, can promote cell migration; B, C are in 22Rv1 and PC3 cell, and shRNA disturbs the expression that reduces CHRM3, reduce the migration of tumor cell.
The specific embodiment
Below in conjunction with accompanying drawing, the specific embodiment provided by the invention is elaborated.
Embodiment
test 1 SABC
Experiment material: normal prostate tissue, prostate cancer tissue; Primary antibodie is the anti-human CHRM3 antibody of rabbit (Abbiotec article No. 251652), the goat anti-rabbit antibody of two anti-HRP labellings (Jackson lab 111-035-003).
Experimental technique:
Organization chip is combined in a large amount of tissue samples a small substrate surface exactly in an orderly manner, by immunohistochemical method, is detected.
1, will organize with the PFA fixative and fix 5 minutes, PBS washes 10 minutes
2, antigen retrieval: with the 0.01M citrate solution, expose epiope.Microwave oven extremely boiling (4 minutes) of heating repair liquid in high fiery 3 minutes, then hang down fiery microwave 1 minute, twice continuously.Be cooled to room temperature, PBS washes 10 minutes.Triton rupture of membranes with 0.5% 10 minutes.
3, sealing nonspecific proteins
1) PBS soaks 3 minutes * 3 times
2) 3% H2O2-methanol (30% H2O210ml+ methanol 90ml) soaking at room temperature is 30 minutes
3) tap water rinses 10 minutes, and PBS soaks 3 minutes * 3 times, and drying or napkin suck unnecessary liquid (not encountering tissue), with the groupization pen, draws circle (distilled water flushing prepares 10% sealing serum in the time) around tissue
4) splashing into rapidly 5% BSA(in circle inner tissue prepares with PBS), sealing heterogenetic antigen (do not allow tissue dry, should add immediately after napkin sops up water), room temperature 30 minutes (preparing primary antibodie), deduct confining liquid, do not wash.
4, primary antibodie is hatched
Get rid of the BSA in section, with filter paper, dry tissue residual BSA on every side, directly add the anti-human CHRM3 antibody (approximately 60 μ l) of rabbit diluted, put into 4 ℃, wet box and spend the night.Second day takes out from refrigerator needs 37 ℃ of rewarming 30 min.
5, two anti-hatching
1) primary antibodie is washed off, slide is inserted to the plastic slide frame, the whole plastic casing of putting into then, add PBS and soak to be put on microoscillator and wash 10 minutes.
2) with filter paper, the water around circle is sucked, add two to resist, room temperature 30 minutes.
3) adding PBS soaks to be put on microoscillator and washes 10 minutes.
6, use the haematoxylin redyeing nucleus, room temperature 30 seconds, clean with tap water.
6, mounting: napkin is absorbed unnecessary liquid, and on slide, dropper drips resinene, and then covered, push gently with tweezers, and drive bubble away, and (tissue site is sure not residual minute bubbles), 60 degree baking boxs are placed 1 hour, Microscopic observation.
test 2 Real-time PCR
Material: prostate cell line, PNT1B, BPH1, C4-2B, DU145, PC3, LNCaP, 22Rv1 etc.
1, (Qianen, Valencia 74134, CA), extract total RNA from prostatic cell to adopt RNeasy Plus mini Kit test kit.Use SYBR Green real-time quantitative RT-PCR technology (Roche, 13962800), detect the expression of CRISP3 gene.Select house-keeping gene GAPDH as internal reference.Real-time quantitative RT-PCR carries out on ABI 7900HT real-time PCR.
2, the expression with the expression standardization CHRM3 gene of internal reference GAPDH by Δ Ct method.With normal prostate cell line PNT1B in contrast.
experiment 3 promotes the tumor cell proliferation experiment
Material: MTS cell proliferation detecting kit (Promega, G358A)
Experimental procedure: get 96 orifice plates, every empty 2,000 cells, volume 100 μ l, when cell increases to 70-80%, add 20 μ l MTS reactant liquors, at 37 ℃, hatches the 2-4 cell, then in microplate reader, detects the absorbance at 490nm place.Because absorbance is directly proportional to cell number and vigor, so can reflect the propagation situation of cell.
PC3 and the 22Rv1 cell for carbachol, processed, after cell attachment, change culture fluid, and the concentration that contains carbachol is followed successively by 1 μ M, 10 μ M and 100 μ M.Continue to cultivate after 24 hours, detect according to the method described above the propagation situation of cell.
test 4 inhibition tumor cell proliferation experiments
Experiment one:
Material: EDU(Invitrogen, C10425)
Experimental procedure: cultured cells is added to EDU, to ultimate density be 10 μ M, 37 degree are 1 hour in the cell culture incubator response time.Then according to the explanation of test kit, add successively reagent.Detect the propagation situation of cell by flow cytometer.
PC3 and the 22Rv1 cell for darifenacin, processed, after cell attachment, change culture fluid, and the concentration that contains darifenacin is followed successively by 1 μ M, 10 μ M, 20 μ M, 50 μ M and 100 μ M.Continue to cultivate after 24 hours, detect according to the method described above the increment situation of cell, cell floating in culture fluid also will be collected.
Experiment two:
In order to check M3 receptor antagonist darifenacin (darifenacin), can suppress in vivo the growth of tumor of prostate, the subcutaneous tumor model that the present invention sets up.
The syringe of material: 1ml, anesthetics (7% chloral hydrate), Matrigel(BD, article No.: 356234)
Experimental technique:
PC3 or 22Rv1 prostate tumor cells are suspended in PBS, 50 microlitres (μ L) are contained to 1, the PBS suspension of 000,000 tumor cell and isopyknic matrigel(matrigel) (article No. 356234) mix homogeneously, and then is injected into the subcutaneous of mice.According to every kg body weight 1mg every day, 5mg or contrast and carry out lumbar injection with normal saline.Measure weekly the size of tumor, the method for calculating is V=a2*b (wherein V is volume, and a is the length of minor face, and b is the length of longest edge).
test 5 tumor migration experiments
Experiment material, the transwell cell of 8 μ m (Corning, 3422), coated with matrigel according to explanation.
Experimental procedure: do transwell when experiment, first by cell in serum-free medium hungry 24 hours, and then the transwell cell above add 100 μ l cell suspension (serum-free), contain 10,000, cell.The complete culture solution that below adds 10% serum of 1ml.In 37 degree cell culture incubators, spend the night.Second day, fix 10 minutes with 4% paraformaldehyde, then uses 0.1% violet staining 10 minutes, after cleaning up, with cotton swab, top cell dabbed off, and after airing, under inverted microscope, observes.
conclusion
Focus on the epicyte protein of the special high expressed of tumor tissues and take for the treatment target spot be a kind ofly be hopeful by it is believed that and effectively find the means for the treatment of the tumor novel drugs.The gene that the genesis of tumor is often expressed with tomour specific is relevant, and the albumen of these tumor cell high expresseds, if function is arranged, may can become the treatment target; If be just again the receptor that is positioned at cell surface, usually can be considered to drug targets, people can produce specific antibody, and can be by antibody and toxin conjugated, special kill tumor cell; Also can adopt agonist or the antagonist of this receptor, promote or suppress receptor active and function, thereby affect growth, propagation and the migration of tumor; If can find this receptor in conjunction with the key factor on albumen or receptor acting signal path, this may be also new biomarker in conjunction with albumen or key factor.For example, VEGF, because rising is expressed in tumor vessel misgrowth, become the active drug for the treatment of clinically tumor.PSMA, the special high expressed albumen of a kind of prostate gland cancer cell, also in clinical trial, and get a good chance of successfully.
M-ChR 3 rich content in prostata tissue; M-ChR 3 is expressed significance than normal prostatic and is raise in prostate cancer tissue; In the outer culture experiment of the body of prostate of neonate rat, add agonist and M3 receptor antagonist can promote respectively and suppress prostatic differentiation.These the results shows, M-ChR 3 can be used as candidate's biomarker for the treatment of novel targets.
M-ChR (Muscarinic Receptors, m receptor) is the acetylcholinergic receptor of G albumen coupling type, in nervous system, plays a significant role.M receptor, by the Gq receptor, can activate phospholipase C, produces IP3 and DAG; Also can promote potassium channels opening or suppress calcium channel; Its β-γ subunit can activate mitogen signal MAPK-ERK, promotes the propagation of cell.The selective agonist research of m receptor is less, and lacks optionally agonist.Cholinergic agonist commonly used has Carbachol, Carbamyl-β-methylcholine chloride, Acetylcholine etc.; M3 receptor-selective antagonist commonly used has Tolterodine, P-F-HHSiD, Oxybutynin, Darifenacin, 4-DAMP etc.The m receptor stirring effect comprises and suppresses circulation, shrinks smooth muscle, dwindles pupil and increase glandular secretion (sweat gland, broncheal gland and digestive gland).M receptor has M1, M2, M3, M4, five kinds of hypotypes of M5, and wherein M4 and M5 mainly are distributed in the central nervous system.M3 receptor high expressed in brain, simultaneously also rich content in prostate.
The present invention finds the treatment target of new tumor of prostate---and M-ChR 3 types (Cholinergic receptor muscarinic 3, CHRM3).Special high expressed M-ChR 3 genes of prostate gland cancer cell and albumen, prove between M-ChR 3 and carcinoma of prostate and have dependency, can be used as the site of prostate targeted therapy.
The present invention confirms that CHRM3 activates propagation and the migration that can promote prostate tumor cells; And, after antagonism CHRM3, can suppress propagation and the migration of prostate tumor cells.
Result of study of the present invention shows: CHRM3 expresses higher than the cancer beside organism of closing on (Figure 1A) in prostate tumor tissue, shows that CHRM3 may play an important role for the genesis of tumor of prostate.The RT-PCR result shows, in prostate tumor cells system, expresses also higher than normal prostate cell line (Figure 1B).
Following the present invention studies CHRM3 in great detail and activates the impact for prostate tumor cells propagation and migration.When adding the stable activator carbachol of CHRM3, we observe the increase of cell PC3 and 22Rv1 propagation.And, cross at CHRM3 the rising (Fig. 2) of also observing cell proliferation in the DU145 of expression and LNCaP cell.Simultaneously, CHRM3 crosses the PNT1B cell of expression, and transfer ability increases, and shows that CHRM3 activates the migration (Fig. 3) that can promote prostate tumor cells.Therefore, we think that the activation of CHRM3 can promote propagation and the migration of prostate tumor cells.In order to confirm to suppress the effect that CHRM3 has the treatment tumor of prostate, we disturb to reduce it by CHRM3 and express in PC3 and 22Rv1 cell, find that the propagation of cell and migration significantly reduce (Fig. 4, Fig. 5), demonstrate the effect of good treatment tumor of prostate.And, there is no at present the target that report can be using the inhibition of CHRM3 as prostate treatment, our discovery may become important clinical treatment target spot, can bring huge social and economical benefits.
Sum up our invention, CHRM3 is high expressed in tumor of prostate, and the activation of CHRM3 can promote propagation and the migration of tumor of prostate, suppresses CHRM3 and can be used as growth and the migration that drug targets suppresses tumor of prostate.
The above is only the preferred embodiment of the present invention; it should be pointed out that for those skilled in the art, under the prerequisite that does not break away from the inventive method; can also make some improvement and supplement, these improvement and supplement and also should be considered as protection scope of the present invention.
Claims (8)
1. the application of M-ChR 3 in the target agent of preparation treatment carcinoma of prostate.
2. application according to claim 1, is characterized in that, the target agent of described treatment carcinoma of prostate is usingd M-ChR 3 as drug targets.
3. M-ChR 3 application in the medicine of screening treatment carcinoma of prostate as target.
4. the application of M-ChR 3, is characterized in that, as the drug targets for the treatment of carcinoma of prostate medicine.
5. the application of M-ChR 3 receptor antagonists in the medicine of preparation treatment carcinoma of prostate.
6. application according to claim 5, is characterized in that, described M-ChR 3 receptor antagonists suppress propagation and the migration of prostate tumor cells.
7. application according to claim 5, is characterized in that, described M-ChR 3 receptor antagonists are Tolterodine, P-F-HHSiD, Oxybutynin, Darifenacin, 4-DAMP.
8. application according to claim 5, is characterized in that, described M-ChR 3 receptor antagonists are Darifenacin.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2013103022648A CN103393628A (en) | 2013-07-18 | 2013-07-18 | Application of muscarinic receptor 3 in treating prostatic cancer |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2013103022648A CN103393628A (en) | 2013-07-18 | 2013-07-18 | Application of muscarinic receptor 3 in treating prostatic cancer |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN103393628A true CN103393628A (en) | 2013-11-20 |
Family
ID=49557490
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2013103022648A Pending CN103393628A (en) | 2013-07-18 | 2013-07-18 | Application of muscarinic receptor 3 in treating prostatic cancer |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103393628A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112237631A (en) * | 2019-07-16 | 2021-01-19 | 清华大学 | Use of acetylcholine pathway modulators in the treatment of cancer |
| WO2024047001A1 (en) | 2022-08-31 | 2024-03-07 | Indivumed Gmbh | Chrm3 as a marker and target for colon cancer therapy |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6642274B1 (en) * | 1999-09-09 | 2003-11-04 | Gary W. Neal | Methods and compositions for preventing and treating prostate disorders |
-
2013
- 2013-07-18 CN CN2013103022648A patent/CN103393628A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6642274B1 (en) * | 1999-09-09 | 2003-11-04 | Gary W. Neal | Methods and compositions for preventing and treating prostate disorders |
Non-Patent Citations (3)
| Title |
|---|
| W.RAYFORD ET AL.: "Muscarinic Cholinergic Receptors Promote Growth of Human Prostate Cancer Cells", 《THE PROSTATE》 * |
| 史一鸣等: "M受体及相关选择性药物研究进展", 《国际药学研究杂志》 * |
| 袁明振等: "前列腺癌和良性前列腺增生组织中M3受体表达及意义", 《中华泌尿外科杂志》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112237631A (en) * | 2019-07-16 | 2021-01-19 | 清华大学 | Use of acetylcholine pathway modulators in the treatment of cancer |
| WO2021008565A1 (en) * | 2019-07-16 | 2021-01-21 | 清华大学 | Use of acetylcholine pathway modulators in treatment of cancer |
| WO2024047001A1 (en) | 2022-08-31 | 2024-03-07 | Indivumed Gmbh | Chrm3 as a marker and target for colon cancer therapy |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Ahmed et al. | Ovarian cancer stem cells: Molecular concepts and relevance as therapeutic targets | |
| Teoh-Fitzgerald et al. | Epigenetic reprogramming governs EcSOD expression during human mammary epithelial cell differentiation, tumorigenesis and metastasis | |
| Elkin et al. | Tail vein assay of cancer metastasis | |
| Lin et al. | Significance of oral cancer‐associated fibroblasts in angiogenesis, lymphangiogenesis, and tumor invasion in oral squamous cell carcinoma | |
| CN102914658A (en) | Application of CRISP3 protein on diagnosis and treatment of prostate cancer | |
| Zhang et al. | Transforming growth factor-β2 is a molecular determinant for site-specific melanoma metastasis in the brain | |
| An et al. | EGFL6 promotes breast cancer by simultaneously enhancing cancer cell metastasis and stimulating tumor angiogenesis | |
| Yang et al. | HIF-2α promotes the formation of vasculogenic mimicry in pancreatic cancer by regulating the binding of Twist1 to the VE-cadherin promoter | |
| Nan et al. | miR-451 suppresses EMT and metastasis in glioma cells | |
| Ruan et al. | CDH11 promotes liver fibrosis via activation of hepatic stellate cells | |
| She et al. | The effect of hepatocellular carcinoma-associated fibroblasts on hepatoma vasculogenic mimicry | |
| CN103966327B (en) | The application of a kind of miR-27a and diagnostic kit thereof | |
| Hao et al. | OPN-a splicing variant expression in non-small cell lung cancer and its effects on the bone metastatic abilities of lung cancer cells in vitro | |
| Lusby et al. | Tumour invasion and dissemination | |
| CN103969452B (en) | The classification diagnosis kit of BAY 43-9006 personalized treatment liver cancer | |
| CN103393628A (en) | Application of muscarinic receptor 3 in treating prostatic cancer | |
| Wu et al. | Expression and significance of c-kit and epithelial-mesenchymal transition (EMT) molecules in thymic epithelial tumors (TETs) | |
| Wang et al. | FGF2 promotes metastasis of uveal melanoma cells via store-operated calcium entry | |
| Fei et al. | Proteomics analysis: inhibiting the expression of P62 protein by chloroquine combined with dacarbazine can reduce the malignant progression of uveal melanoma | |
| CN103911343B (en) | A kind of multiple organ shifts human bladder cancer cell | |
| CN109251982A (en) | The mutant nucleotide sequence of ADGRG6 enhancer, detection method, specific primer used to and application | |
| CN108203734A (en) | Application of the RHCG genes in treating cancer drug and diagnostic kit is prepared | |
| CN107083428A (en) | Applications of the PAK5 in cancer diagnosis prognosis treatment and drug screening | |
| CN106729756A (en) | Application of the biomarker as target in adenocarcinoma of lung diagnosis and treatment | |
| Liu et al. | The role of ethanol in the pathogenesis of non‑bacterial prostatitis |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20131120 |